Stents and catheters are used to facilitate urine drainage within the urinary system. When such sterile implants are inserted into the urinary tract, ions, macromolecules and bacteria from urine, blood or underlying tissues accumulate on their surface. We presented a brief but comprehensive overview of future research strategies in the prevention of urinary device encrustation with an emphasis on biodegradability, molecular, microbiological and physical research approaches. The large and strongly associated field of stent coatings and tissue engineering is outlined elsewhere in this book. There is still plenty of room for future investigations in the fields of material science, surface science, and biomedical engineering to improve and create the most effective urinary implants. In an era where material science, robotics and artificial intelligence have undergone great progress, futuristic ideas may become a reality. These ideas include the creation of multifunctional programmable intelligent urinary implants (core and surface) capable to adapt to the complex biological and physiological environment through sensing or by algorithms from artificial intelligence included in the implant. Urinary implants are at the crossroads of several scientific disciplines, and progress will only be achieved if scientists and physicians collaborate using basic and applied scientific approaches.
\n \n\n \n \nImpaired fracture healing impacts patients\u2019 quality of life and imposes a financial burden on healthcare services. Up to 10% of bone fractures result in delayed/non-union fractures, for which new treatments are urgently required. However, systemic delivery of bone anabolic molecules is often sub-optimal and can lead to significant side effects. In this study, we developed ultrasound (US) responsive nano-sized vehicles in the form of perfluorocarbon nanodroplets (NDs), as a means of targeting delivery of drugs to localised tissues. We tested the hypothesis that NDs could stably encapsulate BIO (GSK-3\u03b2 inhibitor), which could then be released upon US stimulation to activate Wnt signalling and induce ossification. NDs (~280 nm) were prepared from phospholipids and liquid perfluorocarbon and their stability and drug loading was studied by NTA (Nano Tracking Analysis) and HPLC. ND cytotoxicity was assessed in patient-derived bone marrow stromal cells (BMSCs) with Alamar Blue (24 h), and in vitro bioactivity of BIO-NDs was evaluated in a 3T3 Wnt-pathway reporter cell line with luciferase readout. To investigate the acoustic behaviour of NDs, 2% agarose (LM) containing NDs was injected into a bespoke bone fracture model (Sawbones) of various geometries and stimulated by US (1 MHz, 5% duty cycle, 1 MPa, 30 s), allowing the simultaneous capture of optical images and acoustic emissions. Femoral bone hole defects (1\u20132 mm) were made in WT-MF1 mice (age: 8\u201312 wks) and DiR-labelled NDs (100 \u00b5L, 109 NDs/mL, i.v.) were injected post-fracture to determine biodistribution by IVIS imaging. NDs were stable (4 and 37 \u00b0C) and retained >90% BIO until US was applied, which caused ~100% release. ND exposure up to a concentration of 109 NDs/mL showed no cytotoxicity (24 h). BIO-loaded NDs induced Wnt pathway activation in a dose dependent manner. Biodistribution of DiR-NDs in a femoral bone hole defect model in mice demonstrated increased localisation at the fracture site (~2-fold relative to that found in healthy mice or contralateral femurs at 48 h).
\n \n\n \n \nSurfactant-coated gas microbubbles are widely used as contrast agents in ultrasound imaging and increasingly for therapeutic applications. It has been shown that the acoustic response of microbubbles is determined by their size and coating properties and hence depends upon both their chemical composition and the manufacturing technique used to produce them. We have previously presented a hybrid device consisting of a simple microfluidic T-junction with an integrated ultrasound transducer that provides superior production rates and microbubble stability compared with conventional microfluidic systems but with significantly better microbubble uniformity than standard emulsification techniques. The maximum production rate was, however, still limited compared to industrial methods. In the present study a new device was developed that enables production of >108 microbubbles per second using a single device with a mean bubble diameter of 1.4 \u03bcm without degrading microbubble uniformity. Production rates of >109 microbubbles per second can be achieved through parallel operation of multiple channels within a single device; comparable with bulk emulsification but without the risk of contamination and/or degradation of sensitive components.
\n \n\n \n \nImpaired fracture healing is a major financial burden for healthcare services; 5%\u201310% of bone fractures result in non-unions, and there is no clinically approved systemic therapy. This study characterises acoustically stimulated microbubbles (MBs) and nanodroplets (NDs) as non-invasive ultrasound responsive vehicles for the targeted delivery of osteogenic compounds. A microscope-compatible water-tank incorporating a passive cavitation detector was developed to study the acoustic behaviour of MBs and NDs within physical models of bone fractures (gap: 3.5\u20135.5 mm, angles: 0 deg and 90 deg). The device was designed using COMSOL Multiphysics (Burlington, MA) and tested in-vitro. The bone was simulated using a material with comparable acoustic impedance (Sawbones, WA). Numerical simulations showed that the developed experimental set-up generated a relatively uniform acoustic field at a target plane. It could be operated at either 1 or 2 MHz US frequency, at an acoustic pressure in the range 0\u20131 MPa. The inclusion of a fracture model caused perturbations to the acoustic field, which were dependent on the architecture of the fracture (i.e., relative to the incident US field). Ongoing studies are investigating how these perturbations affect ND/MB response in-vitro. Further studies will investigate the relationship between MB/ND acoustic response and the release of biologically active compounds.
\n \n\n \n \nOpportunistic pathogen Pseudomonas aeruginosa plays a crucial role in chronic wound biofilms, increasing infection's morbidity and mortality. In recent years, the signalling molecule nitric oxide (NO) and chelating agent tetrasodium EDTA (T-EDTA) have been applied therapeutically owing to their multifactorial effects including bacterial killing, biofilm dispersal, and wound healing. However, previous studies assessing NO's antibiofilm efficacy have not considered the variable pH and temperature of the wound environment. Here, pH-dependent NO donors N-diazeniumdiolates (NONOates), PAPA NONOate (PA-NO) and Spermine NONOate (SP-NO), and T-EDTA were applied in wound-relevant pH environments (pH 5.5-8.5) and temperatures (32 \u00b0C and 37 \u00b0C) to P. aeruginosa PAO1 biofilms grown for either 24 or 48 h. At 32 \u00b0C and pH 7.5, 250 \u03bcM PA-NO reduced 24-h biofilm biomass by 35 %. At 37 \u00b0C, 250 \u03bcM PA-NO and 4 % w/v T-EDTA caused 21 % and 57 % biomass reduction in 24-h biofilms, respectively. In 48-h biofilms, NONOates did not induce significant biomass reduction, while T-EDTA maintained its efficacy with a 64 % reduction. A subsequent experiment investigated the impact of NONOates and T-EDTA as pre-treatments before exposure to ciprofloxacin. Unexpectedly, NONOate pre-treatment decreased ciprofloxacin's effectiveness, resulting in approximately 1-log increase in viable planktonic and biofilm-residing cells compared to ciprofloxacin alone. It was hypothesized that this protective effect might stem from NO-induced decreased cellular respiration, which inhibits reactive oxygen species (ROS)-mediated bactericidal mechanisms. These findings highlight both the potential and complexities of developing effective antimicrobial strategies for chronic wound infections, emphasizing the need for further research to optimize treatment approaches.
\n \n\n \n \nMicrobubble-enhanced sonothrombolysis is a promising approach to increasing the tolerability and efficacy of current pharmacological treatments for ischemic stroke. Maintaining therapeutic concentrations of microbubbles and drugs at the clot site, however, poses a challenge. The objective of this study was to investigate the effect of magnetic microbubble targeting upon clot lysis rates in vitro. Retracted whole porcine blood clots were placed in a flow phantom of a partially occluded middle cerebral artery. The clots were treated with a combination of tissue plasminogen activator (0.75 \u00b5g/mL), magnetic microbubbles (\u223c107 microbubbles/mL) and ultrasound (0.5 MHz, 630-kPa peak rarefactional pressure, 0.2-Hz pulse repetition frequency, 2% duty cycle). Magnetic targeting was achieved using a single permanent magnet (0.08-0.38 T and 12-140 T/m in the region of the clot). The change in clot diameter was measured optically over the course of the experiment. Magnetic targeting produced a threefold average increase in lysis rates, and linear correlation was observed between lysis rate and total energy of acoustic emissions.
\n \n\n \n \nIn this paper, we propose a simple acoustofluidic method for production of microbubble contrast agents in disposable microfluidic chips by the use of a low-frequency ultrasound transducer and a temperature controller unit. The technique is used to increase the lifetime and to narrow down the size distribution of the microbubbles as compared to traditional microfluidic-based methods.
\n \n\n \n \nNitric oxide (NO) donating drugs such as organic nitrates have been used to treat cardiovascular diseases for more than a century. These donors primarily produce NO systemically. It is however sometimes desirable to control the amount, location, and time of NO delivery. We present the design of a novel pH-sensitive NO release system that is achieved by the synthesis of dipeptide diphenylalanine (FF) and graphene oxide (GO) co-assembled hybrid nanosheets (termed as FF@GO) through weak molecular interactions. These hybrid nanosheets were characterised by using X-ray diffraction, Raman spectroscopy, Fourier transform infrared spectroscopy, zeta potential measurements, X-ray photoelectron spectroscopy, scanning and transmission electron microscopies. The weak molecular interactions, which include electrostatic, hydrogen bonding and \u03c0-\u03c0 stacking, are pH sensitive due to the presence of carboxylic acid and amine functionalities on GO and the dipeptide building blocks. Herein, we demonstrate that this formulation can be loaded with NO gas with the dipeptide acting as an arresting agent to inhibit NO burst release at neutral pH; however, at acidic pH it is capable of releasing NO at the rate of up to 0.6 \u03bcM per minute, comparable to the amount of NO produced by healthy endothelium. In conclusion, the innovative conjugation of dipeptide with graphene can store and release NO gas under physiologically relevant concentrations in a pH-responsive manner. pH responsive NO-releasing organic-inorganic nanohybrids may prove useful for the treatment of cardiovascular diseases and other pathologies.
\n \n\n \n \nBACKGROUND: Ultrasound-responsive microbubbles offer a means of achieving minimally invasive, localised drug delivery in applications including regenerative medicine. To facilitate their use, however, it is important to determine any cytotoxic effects they or their constituents may have. The aim of this study was to test the hypothesis that phospholipid-shelled microbubbles are non-toxic to human bone-derived cells at biologically-relevant concentrations. METHODS: Microbubbles were fabricated using combinations of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dibehenoyl-sn-glycero-3-phosphocholine (DBPC), polyoxyethylene(40) stearate (PEG40S) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene-glycol)-2000] (DSPE-PEG2000). Microbubble size and concentration were measured as a function of time and temperature by optical microscopy. Effects on MG63 osteosarcoma and human bone marrow stromal cells (BMSCs) were measured for up to 72 h by assay for viability, metabolic activity and proliferation. RESULTS: DBPC:DSPE-PEG2000 microbubbles were significantly more stable than DSPC:PEG40S microbubbles under all conditions tested. Serum-containing medium had no detrimental effect on microbubble stability, but storage at 37 \u00b0C compared to at 4 \u00b0C reduced stability for both preparations, with almost complete dissolution of microbubbles at times \u226524 h. DSPC:PEG40S microbubbles had greater inhibitory effects on cell metabolism and growth than DBPC:DSPE-PEG2000 microbubbles, with PEG40S found to be the principle inhibitory component. These effects were only evident at high microbubble concentrations (\u226520% (v/v)) or with prolonged culture (\u226524 h). Increasing cell-microbubble contact by inversion culture in a custom-built device had no inhibitory effect on metabolism. CONCLUSIONS: These data indicate that, over a broad range of concentrations and incubation times, DBPC:DSPE-PEG2000 and DSPC:PEG40S microbubbles have little effect on osteoblastic cell viability and growth, and that PEG40S is the principle inhibitory component in the formulations investigated.
\n \n\n \n \nMicrobubbles utilize high-frequency oscillations under ultrasound stimulation to induce a range of therapeutic effects in cells, often through mechanical stimulation and permeabilization of cells. One of the largest challenges remaining in the field is the characterization of interactions between cells and microbubbles at therapeutically relevant frequencies. Technical limitations, such as employing sufficient frame rates and obtaining sufficient image resolution, restrict the quantification of the cell's mechanical response to oscillating microbubbles. Here, a novel methodology was developed to address many of these limitations and improve the image resolution of cell-microbubble interactions at high frame rates. A compact acoustic device was designed to house cells and microbubbles as well as a therapeutically relevant acoustic field while being compatible with a Shimadzu HPV-X camera. Cell viability tests confirmed the successful culture and proliferation of cells, and the attachment of DSPC- and cationic DSEPC-microbubbles to osteosarcoma cells was quantified. Microbubble oscillation was observed within the device at a frame rate of 5 million FPS, confirming suitable acoustic field generation and ultra high-speed image capture. High spatial resolution in these images revealed observable deformation in cells following microbubble oscillation and supported the first use of digital image correlation for strain quantification in a single cell. The novel acoustic device provided a simple, effective method for improving the spatial resolution of cell-microbubble interaction images, presenting the opportunity to develop an understanding of the mechanisms driving the therapeutic effects of oscillating microbubbles upon ultrasound exposure.
\n \n\n \n \nBacterial biofilms are an ever-growing concern for public health, featuring both inherited genetic resistance and a conferred innate tolerance to traditional antibiotic therapies. Consequently, there is a growing interest in novel methods of drug delivery, in order to increase the efficacy of antimicrobial agents. One such method is the use of acoustically activated microbubbles, which undergo volumetric oscillations and collapse upon exposure to an ultrasound field. This facilitates physical perturbation of the biofilm and provides the means to control drug delivery both temporally and spatially. In line with current literature in this area, this review offers a rounded argument for why ultrasound-responsive agents could be an integral part of advancing wound care. To achieve this, we will outline the development and clinical significance of biofilms in the context of chronic infections. We will then discuss current practices used in combating biofilms in chronic wounds and then critically evaluate the use of acoustically activated gas microbubbles as an emerging treatment modality. Moreover, we will introduce the novel concept of microbubbles carrying biologically active gases that may facilitate biofilm dispersal.
\n \n\n \n \nObstructions of the ureter lumen can originate from intrinsic or extrinsic factors, such as kidney stones, tumours, or strictures. These can affect the physiological flow of urine from the kidneys to the bladder, potentially causing infection, pain, and kidney failure. To overcome these complications, ureteral stents are often deployed clinically in order to temporarily re-establish urinary flow. Despite their clinical benefits, stents are prone to encrustation and biofilm formation that lead to reduced quality of life for patients; however, the mechanisms underlying the formation of crystalline biofilms in stents are not yet fully understood. In this study, we developed microfluidic-based devices replicating the urodynamic field within different configurations of an occluded and stented ureter. We employed computational fluid dynamic simulations to characterise the flow dynamic field within these models and investigated bacterial attachment (Pseudomonas fluorescens) by means of crystal violet staining and fluorescence microscopy. We identified the presence of hydrodynamic cavities in the vicinity of a ureteric occlusion, which were characterised by low levels of wall shear stress (WSS < 40 mPa), and observed that initiation of bacterial attachment occurred in these specific regions of the stented ureter. Notably, the bacterial coverage area was directly proportional to the number of cavities present in the model. Fluorescence microscopy confirmed that the number density of bacteria was greater within cavities (3 bacteria\u00b7mm-2) when compared to side-holes of the stent (1 bacterium\u00b7mm-2) or its luminal surface (0.12\u00b7bacteria mm-2). These findings informed the design of a novel technological solution against bacterial attachment, which reduces the extent of cavity flow and increases wall shear stress over the stent's surface.
\n \n\n \n \nINTRODUCTION: An ideal stent would offer simple insertion and removal with no discomfort and/or migration, it would have no biofilm formation or encrustation and would also maintain the patient's quality of life. MATERIAL AND METHODS: In this mini-review, we outlined the engineering developments related to stent material, design and coating. RESULTS: There have been a wide variety of in-vitro, model-based, animal-based and clinical studies using a range of commercial and non-commercial stents. Ureteric stents have evolved since their first usage with a wider range of stent design, material and coating available for laboratory and clinical use. CONCLUSIONS: While engineering innovations have led to the evolution of stents, more work needs to be done to address the issues relating to stent encrustation and biofilm formation.
\n \n\n \n \nWe demonstrate an imaging flow cytometer that uses acoustic levitation to assemble cells and other particles into a sheet structure. This technique enables a high resolution, low noise CMOS camera to capture images of thousands of cells with each frame. While ultrasonic focussing has previously been demonstrated for 1D cytometry systems, extending the technology to a planar, much higher throughput format and integrating imaging is non-trivial, and represents a significant jump forward in capability, leading to diagnostic possibilities not achievable with current systems. A galvo mirror is used to track the images of the moving cells permitting exposure times of 10 ms at frame rates of 50 fps with motion blur of only a few pixels. At 80 fps, we demonstrate a throughput of 208\u2009000 beads per second. We investigate the factors affecting motion blur and throughput, and demonstrate the system with fluorescent beads, leukaemia cells and a chondrocyte cell line. Cells require more time to reach the acoustic focus than beads, resulting in lower throughputs; however a longer device would remove this constraint.
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